The cellular expression of immune response (Ir) gene function was studied in both primary and secondary in vitro antibody responses to the TNP conjugates of (T,G)-A--L and (H,G)-A--L. It was demonstrated that the function of accessory cells in responses to TNP-(T,G)-A--L and TNP-(H,G)-A--L is under the control of genes which also map to K or I-A. In contrast, the expression of Ir gene function by B cells is related to the B cell activation pathway; Ir gene function is expressed by Lyb5- B cells activated under appropriate conditions, while Ir genes do not influence Lyb5+ B cell function under different conditions. In vitro augmented primary and secondary responses to TNP-nuclease (TNP-NASE) have also been established and documented to be under the control of H-2 linked Ir gene(s) mapping to the I-B subregion. For these responses, accessory cell function was shown to be under Ir gene control. Recent data employing monoclonal anti-Ia reagents have suggested that genes in the I-A subregion may also be involved in regulating responses to TNP-NASE.